Autoimmunities
Abstract
Therapeutic strategies targeting complement have revolutionized the treatment of myasthenia gravis (MG). However, a deeper understanding of complement modulation in the human system is required to improve treatment responses and identify “off-target effects” shaping long-term outcomes. For this purpose, we studied a cohort of MG patients treated with either eculizumab (n = 10) or azathioprine (n = 10) as well as treatment-naïve (n = 10) patients using a combined proteomics and metabolomics approach. This strategy confirmed known effects of eculizumab on the terminal complement cascade. Beyond that, eculizumab modulated the serum proteometabolome as distinct pathways were altered in eculizumab-treated patients including the oxidative stress response, mitogen-activated protein kinase signaling and lipid metabolism with particular emphasis on arachidonic acid signaling. We detected reduced levels of arachidonate 5-lipoxygenase (ALOX5) and leukotriene A4 (LTA4) in eculizumab-treated patients. Mechanistically, ligation of the C5a receptor (C5aR) is needed for ALOX5 metabolism and generation of downstream leukotrienes. As eculizumab prevents cleavage of C5 into C5a, decreased engagement of C5aR may inhibit ALOX5-mediated synthesis of pro-inflammatory leukotrienes. These findings indicate distinct “off-target effects” induced by eculizumab, illuminating potential mechanisms of action that may be harnessed to improve treatment outcomes.
Authors
Christopher Nelke, Christina B. Schroeter, Frauke Stascheit, Niklas Huntemann, Marc Pawlitzki, Alice G. Willison, Saskia Räuber, Nico Melzer, Ute Distler, Stefan Tenzer, Kai Stühler, Andreas Roos, Andreas Meisel, Sven G. Meuth, Tobias Ruck
Abstract
Methotrexate (MTX) is a standard, first-line therapy for rheumatoid arthritis (RA); however, its precise mechanisms of action other than antifolate activity are largely unknown. We performed DNA microarray analyses of CD4+ T cells in patients with RA before and after MTX treatment and found that TP63 was the most significantly downregulated gene after MTX treatment. TAp63, an isoform of TP63, was highly expressed in human IL-17–producing Th (Th17) cells and was suppressed by MTX in vitro. Murine TAp63 was expressed at high levels in Th cells and at lower levels in thymus-derived Treg cells. Importantly, TAp63 knockdown in murine Th17 cells ameliorated the adoptive transfer arthritis model. RNA-Seq analyses of human Th17 cells overexpressing TAp63 and those with TAp63 knockdown identified FOXP3 as a possible TAp63 target gene. TAp63 knockdown in CD4+ T cells cultured under Th17 conditions with low-dose IL-6 increased Foxp3 expression, suggesting that TAp63 balances Th17 cells and Treg cells. Mechanistically, TAp63 knockdown in murine induced Treg (iTreg) cells promoted hypomethylation of conserved noncoding sequence 2 (CNS2) of the Foxp3 gene and enhanced the suppressive function of iTreg cells. Reporter analyses revealed that TAp63 suppressed the activation of the Foxp3 CNS2 enhancer. Collectively, TAp63 suppresses Foxp3 expression and exacerbates autoimmune arthritis.
Authors
Kensuke Suga, Akira Suto, Shigeru Tanaka, Yutaka Sugawara, Takahiro Kageyama, Junichi Ishikawa, Yoshie Sanayama, Kei Ikeda, Shunsuke Furuta, Shin-Ichiro Kagami, Arifumi Iwata, Koichi Hirose, Kotaro Suzuki, Osamu Ohara, Hiroshi Nakajima
Abstract
Myelin oligodendrocyte glycoprotein (MOG) antibody-associated disease (MOGAD) is an inflammatory demyelinating central nervous system condition characterized by the presence of MOG autoantibodies. We sought to investigate whether human MOG autoantibodies are capable of mediating damage to MOG-expressing cells through multiple mechanisms. We developed high-throughput assays to measure complement activity (CA), complement-dependent cytotoxicity (CDC), antibody-dependent cellular phagocytosis (ADCP), and antibody-dependent cellular cytotoxicity (ADCC) of live MOG-expressing cells. MOGAD patient sera effectively mediate all of these effector functions. Our collective analyses reveal that [i] cytotoxicity is not incumbent on MOG autoantibody quantity alone, [ii] engagement of effector functions by MOGAD patient serum is bimodal, with some sera exhibiting cytotoxic capacity while others did not, [iii] the magnitude of CDC and ADCP is elevated closer to relapse, while MOG-IgG binding is not, and [iv] all IgG subclasses can damage MOG-expressing cells. Histopathology from a representative MOGAD case revealed congruence between lesion histology and serum CDC and ADCP, and we identified NK cells, mediators of ADCC, in the cerebrospinal fluid of relapsing MOGAD patients. Thus, MOGAD-derived autoantibodies are cytotoxic to MOG-expressing cells through multiple mechanisms and assays quantifying CDC and ADCP may prove to be effective tools for predicting risk of future relapses.
Authors
Soumya S. Yandamuri, Beata Filipek, Abeer H. Obaid, Nikhil Lele, Joshua M. Thurman, Naila Makhani, Richard J. Nowak, Yong Guo, Claudia F. Lucchinetti, Eoin P. Flanagan, Erin E. Longbrake, Kevin C. O'Connor
Abstract
Abdominal aortic aneurysm (AAA) is usually asymptomatic until life-threatening complications occur, predominantly involving aortic rupture. Currently, no drug-based treatments are available, primarily due to limited understanding of AAA pathogenesis. Transcriptional regulator PR domain-containing protein 16 (PRDM16) is highly expressed in the aorta, but its functions in the aorta are largely unknown. By RNA-seq analysis, we found that VSMCs-specific Prdm16 knockout mice (Prdm16SMKO) already showed extensive changes in the expression of genes associated with extracellular matrix (ECM) remodeling and inflammation in the abdominal aorta under normal housing conditions without any pathological stimuli. Human AAA lesions displayed lower PRDM16 expression. Periadventitial elastase application to the suprarenal region of the abdominal aorta aggravated AAA formation in Prdm16SMKO. During AAA development, VSMCs undergo apoptosis because of both intrinsic and environmental changes including inflammation and ECM remodeling. Prdm16 deficiency promoted inflammation and apoptosis in VSMCs. A disintegrin and metalloproteinase 12 (ADAM12) is a gelatinase which can degrade various ECM. We found that ADAM12 is a target of transcriptional repression by PRDM16. Adam12 knockdown reversed VSMC apoptosis induced by Prdm16 deficiency. Our study demonstrated that PRDM16 deficiency in VSMCs promoted ADAM12 expression and aggravates AAA formation, which may provide potential targets for AAA treatment.
Authors
Zhenguo Wang, Xiangjie Zhao, Guizhen Zhao, Yanhong Guo, Haocheng Lu, Wenjuan Mu, Juan Zhong, Minerva Garcia-Barrio, Jifeng Zhang, Y. Eugene Chen, Lin Chang
Abstract
Primary Sjogren’s syndrome (pSS) is a systemic autoimmune inflammatory disease mainly defined by T cell–dominated destruction of exocrine glands. Currently, CD8+T cells were closely related to the pathogenesis of pSS. However, the single-cell immune profiling of pSS and molecular signatures of pathogenic CD8+T cells have not been well elucidated. Our multiomics investigation identified that both T cell and B cell, especially CD8+T cells, were undergoing significant clonal expansion in pSS patients. TCR clonality analysis revealed that peripheral granzyme (GZM) K+CXCR6+CD8+T cells had higher proportion of shared clones with CD69+CD103-CD8+ tissue resident memory T (TRM) cells in labial glands in pSS. CD69+CD103-CD8+TRM cells featured by high expression of GZMK were more active and cytotoxic in pSS compared with their CD103+ counterparts. Peripheral GZMK+CXCR6+CD8+T cells with higher CD122 expression were increased and harbored a gene signature similar to TRM cells in pSS. Consistently, IL-15 was significantly elevated in pSS plasma and showed the capacity to promote differentiation of CD8+T cells into GZMK+CXCR6+CD8+T cells in a STAT5 dependent manner. Taken together, we depicted the immune landscape of pSS and further conducted comprehensive bioinformatics analysis and in vitro experimental investigation to characterize the pathogenic role and differentiation trajectory of CD8+TRM cells in pSS.
Authors
Ting Xu, Hao-Xian Zhu, Xing You, Jin-Fen Ma, Xin Li, Pan-Yue Luo, Yang Li, Zhe-Xiong Lian, Cai-Yue Gao
Abstract
Low capacity to produce reactive oxygen species (ROS) due to mutations in neutrophil cytosolic factor 1 (NCF1/p47phox), a component of NADPH oxidase 2 (NOX2) complex, is strongly associated with systemic lupus erythematosus in both humans and mouse models. Here, we aim to identify the key immune cell type(s) and cellular mechanisms driving lupus pathogenesis under the condition of NCF1-dependent ROS deficiency. Using a set of cell-specific Cre-deleter, the human NCF1-339 variant knock-in, and transgenic mouse strains, we show that low ROS production in plasmacytoid dendritic cells (pDCs) exacerbates both pristane-induced lupus and a newly established Yaa-related spontaneous model by promoting pDC accumulation in multiple organs during lupus development, accompanied by elevated IFNα levels and expression of IFN-stimulated genes. Mechanistic studies reveal that ROS deficiency enhances pDC generation through the AKT/mTOR pathway and CCR2-mediated migration to tissues, which together with hyperactivation of the redox-sensitive STING/IFNα/JAK1/STAT1 cascade further augments type I IFN responses. More importantly, by suppressing these pathways, restoration of NOX2-derived ROS specifically in pDCs protects against lupus. These discoveries explain the causative effect of dysfunctional NCF1 in lupus and demonstrate the protective role of pDC-derived ROS in disease development driven by NCF1-dependent ROS deficiency.
Authors
Huqiao Luo, Vilma Urbonaviciute, Amir Ata Saei, Hezheng Lyu, Massimiliano Gaetani, Ákos Végvári, Yanpeng Li, Roman A. Zubarev, Rikard Holmdahl
Abstract
The widespread presence of autoantibodies in acute infection with SARS-CoV-2 is increasingly recognized, but the prevalence of autoantibodies in non–SARS-CoV-2 infections and critical illness has not yet been reported. We profiled IgG autoantibodies in 267 patients from 5 independent cohorts with non–SARS-CoV-2 viral, bacterial, and noninfectious critical illness. Serum samples were screened using Luminex arrays that included 58 cytokines and 55 autoantigens, many of which are associated with connective tissue diseases (CTDs). Samples positive for anti-cytokine antibodies were tested for receptor blocking activity using cell-based functional assays. Anti-cytokine antibodies were identified in > 50% of patients across all 5 acutely ill cohorts. In critically ill patients, anti-cytokine antibodies were far more common in infected versus uninfected patients. In cell-based functional assays, 11 of 39 samples positive for select anti-cytokine antibodies displayed receptor blocking activity against surface receptors for Type I IFN, GM-CSF, and IL-6. Autoantibodies against CTD-associated autoantigens were also commonly observed, including newly detected antibodies that emerged in longitudinal samples. These findings demonstrate that anti-cytokine and autoantibodies are common across different viral and nonviral infections and range in severity of illness.
Authors
Allan Feng, Emily Y. Yang, Andrew Reese Moore, Shaurya Dhingra, Sarah Esther Chang, Xihui Yin, Ruoxi Pi, Elisabeth K.M. Mack, Sara Völkel, Reinhard Geßner, Margrit Gündisch, Andreas Neubauer, Harald Renz, Sotirios Tsiodras, Paraskevi C. Fragkou, Adijat A. Asuni, Joseph E. Levitt, Jennifer G. Wilson, Michelle Leong, Jennifer H. Lumb, Rong Mao, Kassandra Pinedo, Jonasel Roque, Christopher M. Richards, Mikayla Stabile, Gayathri Swaminathan, Maria L. Salagianni, Vasiliki Triantafyllia, Wilhelm Bertrams, Catherine A. Blish, Jan E. Carette, Jennifer Frankovich, Eric Meffre, Kari Christine Nadeau, Upinder Singh, Taia T. Wang, Eline T. Luning Prak, Susanne Herold, Evangelos Andreakos, Bernd Schmeck, Chrysanthi Skevaki, Angela J. Rogers, Paul J. Utz
Abstract
Neuromyelitis optica spectrum disorders (NMOSD) are inflammatory autoimmune disorders of the CNS. Immunoglobulin G autoantibodies targeting the aquaporin-4 water channel (AQP4-IgG) are the pathogenic effector of NMOSD. Dysregulated T follicular helper (Tfh) cells have been implicated in the loss of B-cell tolerance in autoimmune diseases. The contribution of Tfh cells to disease activity and the therapeutic potential of targeting these cells in NMOSD remain unclear. Here, we established an autoimmune model of NMOSD by immunizing mice against AQP4 via in vivo electroporation. After AQP4 immunization, mice displayed AQP4 autoantibodies in the blood circulation, blood-brain barrier disruption, and IgG infiltration in the spinal cord parenchyma. Moreover, AQP4 immunization induced motor impairments and NMOSD-like pathologies including astrocytopathy, demyelination, axonal loss, and microglia activation. These were associated with increased splenic Tfh, T helper 1 (Th1) and T helper 17 (Th17) cells, memory B cells and plasma cell. AQP4-deficient mice did not displayed motor impairments and NMOSD-like pathologies after AQP4 immunization. Importantly, abrogating inducible costimulator (ICOS)/inducible costimulator ligand (ICOS-L) signalling using anti-ICOS-L antibody depleted Tfh cells and suppressed the response of Th1 and Th17 cells, memory B cells, and plasma cells in AQP4-immunized mice. These findings were associated with ameliorated motor impairments and spinal cord pathologies. This study suggests a role of Tfh cells in the pathophysiology of NMOSD in a novel mouse model with AQP4 autoimmunity. It also provides an animal model for further investigating the immunological mechanisms underlying AQP4 autoimmunity, and for developing novel therapeutic interventions targeting the autoimmune reactions in NMOSD.
Authors
Leung-Wah Yick, Oscar Ka-Fai Ma, Ethel Yin-Ying Chan, Krystal Xiwing Yau, Jason Shing-Cheong Kwan, Koon-Ho Chan
Abstract
A GWAS of patients with anti-neutrophil cytoplasmic antibodies (ANCA) found an association between proteinase-3 (PR3) ANCA and a single-nucleotide polymorphism (SNP) (rs62132293) upstream of PRTN3, encoding PR3. The variant (G-allele) was shown to be an eQTL in healthy controls, but the clinical impact remains unknown. Longitudinally followed ANCA patients(n=401) and healthy controls(n=130) were genotyped. Gene expression was quantified by RT-qPCR from leukocyte RNA. Plasma PR3 was quantified by ELISA. Kaplan-Meier estimates and log rank test were used for clinical outcomes. Among patients, variant carriers had elevated leukocyte PRTN3 expression compared to non-carriers (C/G vs. C/C and G/G vs. C/C, p=0.012 and p=0.001, respectively, effect size 0.24). Healthy controls had low PRTN3 regardless of genotype. MPO expression did not differ by genotype. PRTN3 (message) correlated with circulating PR3 (r=0.36, p<0.0005) and variant carriers had higher plasma PR3 compared to non-carriers (p=0.041). Among variant carriers, there was a 1.66-fold increased risk of relapse in patients with PR3-ANCA vs MPO-ANCA (HR 1.66, 95% CI 1.08, 2.54). The risk allele marked by rs62132293 is clinically significant as it is associated with increased autoantigen and may, in part, explain increased relapse in PR3-ANCA. Our results underscore the role of autoantigen availability in ANCA vasculitis.
Authors
Dhruti P. Chen, Claudia P. Aiello, DeMoris A. McCoy, Taylor Stamey, Jiajin Yang, Susan L. Hogan, Yichun Hu, Vimal K. Derebail, Eveline Y. Wu, J. Charles Jennette, Ronald J. Falk, Dominic J. Ciavatta
Abstract
CD4+ cytotoxic T lymphocytes (CTLs) were recently implicated in immune-mediated inflammation and fibrosis progression of Graves’ orbitopathy (GO). However, little is known about therapeutic targeting CD4+ CTLs. Herein, we studied the effect of rapamycin, an approved mTORC1 inhibitor, in GO mouse model, in vitro and in refractory GO patients. In the adenovirus-induced model, rapamycin significantly decreased the incidence of orbitopathy. This was accompanied by reduction of CD4+ CTLs, and improvement of inflammation, adipogenesis and fibrosis in orbits. CD4+CTLs from active GO patients showed upregulation of mTOR pathway, while rapamycin decreased their proportions and cytotoxic function. Low-dose rapamycin treatment substantially improved diplopia and clinical activity score in steroid-refractory GO patients. Single-cell RNA sequencing revealed that eye motility improvement was closely related to suppression of inflammation and chemotaxis in CD4+ CTLs. In conclusion, rapamycin is a promising treatment for CD4+ CTL-mediated inflammation and fibrosis in GO.
Authors
Meng Zhang, Kelvin K.L. Chong, Zi-yi Chen, Hui Guo, Yu-feng Liu, Yong-yong Kang, Yang-jun Li, Ting-ting Shi, Kenneth Ka Hei Lai, Ming-qian He, Kai Ye, George J. Kahaly, Bing-yin Shi, Yue Wang
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